The present invention relates to novel proteins which induce death of cancer cells by inducing apoptosis of the cells, or which exhibit cancer cell growth inhibitory activity, genes of such proteins, monoclonal antibodies to such proteins, reagents for qualifying cancer cell apoptosis, and carcinostatic agents.
Remarkable development in chemotherapy has been improving survival rate and remedial rate of patients having neoplastic diseases. On the other hand, however, strong side effects of carcinostatic agents give serious damage on normal cells, which has been recognized as a social problem. For prevention of side effects of carcinostatic agents, agents are demanded which have excellent selectivity for cancer cells or which are capable of controlling proliferation of oncocytes.
In conventional chemotherapy for cancer, administration dosage of agents is kept as low as possible since even the treatment dosage may cause harmful effects. Thus, attempts have been made to seek for potentiation by combination of a plurality of carcinostatic agents with different mechanisms of actions, or to improve carcinostatic effect by combination of a carcinostatic agent with other substances. In the latter case, a carcinostatic agent is usually combined with an immune activator to combine the direct effect on oncocytes with antineoplastic effect obtained through activation of immunocompetence of organism. Further, in some cases, radiotherapy or surgical treatment is performed in addition to these methods to improve the effect of the treatment.
Recently, it has been revealed that there are two different types of cell death, i.e., apoptosis (cell death governed by genes) and necrosis (cell death not governed by genes). Apoptosis and necrosis are usually distinguished by observing DNA fragmentation through biochemical measurement. This measurement shows that conventional carcinostatic agents exhibit carcinostatic effect by inducing necrosis of oncocytes, so that they cannot genetically control the death of oncocytes. On the contrary, active researches have been made on substances which induce apoptosis of oncocytes since such substances have possibility to control the death of oncocytes. Combination of an apoptosis-inducing carcinostatic agent with a conventional carcinostatic agent having different mechanism of action is expected to improve antineoplastic effect.
It is an object of the present invention to provide novel proteins which induce death of cells, in particular human cancer cells, by inducing apoptosis of the cells, or which exhibits cancer cell growth inhibitory activity; fragments of such proteins; genes of such proteins; and monoclonal antibodies to such protein.
It is another object of the present invention to provide reagents for apoptosis induction which induce apoptosis of cells, in particular of cancer cells, in vitro for the study of mechanism of cell death induction.
It is still another object of the present invention to provide carcinostatic agents which have proliferation inhibitory effect on human cancer cells or which have death inducing effect on cancer cells.
The present inventors have conducted intensive researches on peptides or proteins which induce apoptosis of cancer cells for applying conventionally known apoptotic cell death to oncotherapy. As a result, the inventors have found that some proteins purified from mackerel""s viscus have apoptosis-inducing activity not only on blood cancer cells but also on a variety of tumor cancer cells, then synthesized cDNA, using the mRNA isolated from mackerel""s viscus, determined the DNA sequence of the obtained cDNA, and estimated the amino acid sequence thereof, thereby completing the present invention.
In sum, according to the present invention, there are provided proteins comprising the amino acid sequence of SEQ ID No. 1 of the attached sequence listing, or the amino acid sequence at least partly homologous or analogous to the sequence of SEQ ID No. 1, and having proliferation inhibitory activity on cancer cells or cell death inducing activity; and fragments of such proteins.
According to the present invention, there are also provided proteins which have the amino acid sequence homologous to the amino acid sequence of SEQ ID No. 1 of the attached sequence listing, and which have proliferation inhibitory activity on cancer cells or cell death inducing activity.
According to the present invention, there are also provided proteins comprising amino acids 61-89 and 497-514 of SEQ ID No. 1 of the attached sequence listing, and having proliferation inhibitory activity on cancer cells or cell death inducing activity.
According to the present invention, there are further provided genes which code the aforementioned proteins or their fragments, and genes which code the aforementioned proteins or their fragments and which comprise the DNA sequence of SEQ ID No. 2 of the attached sequence listing or the DNA sequence at least partly homologous or analogous to that of SEQ ID No. 2.
According to the present invention, there are further provided monoclonal antibodies to the aforementioned proteins or their fragments.
According to the present invention, there are also provided reagents for qualifying apoptosis comprising at least one member selected from the group consisting of the aforementioned proteins, their fragments, and the monoclonal antibodies thereto.
According to the present invention, there are also provided carcinostatic agents comprising the aforementioned proteins and their fragments as active components.